A locking mechanism preventing radical damage in the absence of substrate, as revealed by the x-ray structure of lysine 5,6-aminomutase

doi:10.1073/pnas.0407074101

A locking mechanism preventing radical damage in the absence of substrate, as revealed by the x-ray structure of lysine 5,6-aminomutase

^*Department of Chemistry, Massachusetts Institute of Technology, 77 Massachusetts Avenue, Cambridge, MA 02139; and ^†Department of Biochemistry, University of Wisconsin, 1710 University Avenue, Madison, WI 53726

Contributed by Perry A. Frey, September 24, 2004

Abstract

Lysine 5,6-aminomutase is an adenosylcobalamin and pyridoxal-5′-phosphate-dependent enzyme that catalyzes a 1,2 rearrangement of the terminal amino group of dl-lysine and of l-β-lysine. We have solved the x-ray structure of a substrate-free form of lysine-5,6-aminomutase from Clostridium sticklandii. In this structure, a Rossmann domain covalently binds pyridoxal-5′-phosphate by means of lysine 144 and positions it into the putative active site of a neighboring triosephosphate isomerase barrel domain, while simultaneously positioning the other cofactor, adenosylcobalamin, ≈25 Å from the active site. In this mode of pyridoxal-5′-phosphate binding, the cofactor acts as an anchor, tethering the separate polypeptide chain of the Rossmann domain to the triosephosphate isomerase barrel domain. Upon substrate binding and transaldimination of the lysine-144 linkage, the Rossmann domain would be free to rotate and bring adenosylcobalamin, pyridoxal-5′-phosphate, and substrate into proximity. Thus, the structure embodies a locking mechanism to keep the adenosylcobalamin out of the active site and prevent radical generation in the absence of substrate.

Footnotes

↵ § To whom correspondence should be addressed at: Department of Chemistry, 16-573, Massachusetts Institute of Technology, Cambridge, MA 02139. E-mail: cdrennan{at}mit.edu.
↵ ‡ Present address: Department of Biochemistry, Beckman Center B400, Stanford University, Stanford, CA 94305-5307.
Author contributions: E.B., P.A.F., and C.L.D. designed research; F.B. and E.A.E. performed research; E.B., K.-H.T., and P.A.F. contributed new reagents/analytic tools; F.B. and C.L.D. analyzed data; and F.B. and C.L.D. wrote the paper.
Abbreviations: AdoCbl, adenosylcobalamin or coenzyme B₁₂; Ado, adenosyl group; Ado^•, 5′-deoxyadenosyl radical; AdoH, 5′-deoxyadenosine; AdoMet, S-adenosylmethionine; Cbl, cobalamin; DMB, dimethylbenzimidazole; GM, glutamate mutase; 5,6-LAM, lysine 5,6-aminomutase; 2,3-LAM, lysine 2,3-aminomutase; MCM, methylmalonyl-coenzyme A mutase; PLP, pyridoxal 5′-phosphate; TIM, triosephosphate isomerase.
Data deposition: The atomic coordinates and structure factors have been deposited in the Protein Data Bank, www.pdb.org (PDB ID code 1XRS).